Nano-drug transdermal delivery system as well as preparation method and application thereof

Abstract

The invention provides a nano-drug transdermal delivery system as well as a preparation method and application thereof. According to the nano-drug transdermal delivery system, lipoic acid is taken as a drug carrier, a chemotherapy drug and / or a photodynamic reagent are / is entrapped, and nano-particles containing 1, 2-dithiolane are formed through self-assembly. According to the invention, the lipoic acid is used as a carrier to assist micromolecules in a transdermal delivery system for the first time, no extra transdermal penetration enhancer is needed, the control of skin position and accessory number is not needed, the auxiliary administration of equipment is not needed, and effective transdermal delivery can be realized only by simple smearing. The administration mode is non-invasive, efficient transdermal penetration can be realized without destroying cuticle, and the risk of skin infection is reduced. The transdermal depth can reach more than 1mm, and the skin care product is suitable for superficial and deep skin diseases. Autonomous operation can be achieved, cost is remarkably reduced, and safety is high. On the basis, the types of medicines to be loaded can be flexibly changed according to the types of skin diseases, and more accurate treatment is realized.

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical technology, and in particular to a nano drug transdermal delivery system and a preparation method and application thereof. Background Art

[0002] As the largest organ in the human body, the skin has many functions, such as regulating body temperature, preventing the loss of water and salt, and resisting the invasion of various pathogens, microorganisms and viruses. The skin structure is divided into two layers, the outermost epidermis and the inner dermis. The skin layers are subject to endogenous gene regulation or various exogenous stimuli, such as gene mutation, ultraviolet radiation, trauma, etc., which can cause various diseases. The occurrence of these diseases will seriously reduce people's quality of life, such as scar hyperplasia, psoriasis, psoriasis, etc. Some are even life-threatening, such as basal carcinoma, squamous cell carcinoma, melanoma and other skin cancers. Therefore, it is necessary to develop various effective treatment methods to improve the treatment efficiency of skin diseases.

[0003] Drug delivery through the skin has many advantages, including increasing patient compliance, avoiding the first-pass effect of oral drugs on the liver, achieving long-term controlled release of drugs, and reducing the need for multiple doses.

[0004] Although transdermal drug delivery has unique advantages, most drugs cannot effectively penetrate the dense stratum corneum, the outermost layer of the skin. Preparation of transdermal preparations generally requires that the molecular weight of the drug should be less than 500Da, otherwise it is difficult to penetrate the stratum corneum. The solubility is greater than 1mg / mL, and the absolute value of the oil-water partition coefficient is 1-2, so as to ensure that the drug can both penetrate the stratum corneum and enter the active epidermis. Due to these restrictions, the types of drugs that can be used for transdermal drug delivery are very few.

[0005] Among them, nano drug delivery systems are widely reported in the field of transdermal delivery due to their unique physicochemical properties. Compared with other transdermal drug delivery systems (TDDS), nano drugs generally show better drug release, deeper drug penetration, and allow encapsulation of hydrophilic and hydrophobic drug molecules. Nano drugs generally achieve effective transdermal delivery through three pathways: intercellular lipid pathway, transcellular pathway, and hair follicle pathway. Generally, fat-soluble and amphiphilic molecules pass through the intercellular lipid pathway, but the permeability of this pathway is low, and penetration enhancers need to be added to improve the efficiency of transdermal delivery, which can irritate the skin and cause allergies, inflammation, etc. The transcellular pathway requires nano drugs to alternate between hydrophilic (inside the cell) and lipophilic areas (extracellular matrix). Due to the continuous diffusion and distribution between these areas, this pathway is not conducive to most drugs. Nano drugs can also penetrate through skin appendages such as hair follicles, sebaceous glands, and sweat glands. However, these skin appendages only account for 0.1%-1% of the skin surface area, and the delivery efficiency is limited. In addition, the density of skin appendages varies in different skin regions, limiting the application sites for transdermal delivery of nanomedicines, which reduces the types of diseases that can be treated.

[0006] Therefore, developing a new and efficient transdermal drug administration route and a non-invasive transdermal delivery system that is safe and not affected by the physicochemical properties of the drug and the skin condition has important practical significance for clinical treatment. Summary of the invention

[0007] In view of this, the technical problem to be solved by the present invention is to provide a nano drug transdermal delivery system and a preparation method and application thereof, wherein the nano drug can deliver drugs through a thiol exchange pathway for single treatment or combined treatment of skin diseases.

[0008] To achieve the above object, the present invention provides the use of lipoic acid as a transdermal nano drug carrier.

[0009] The lipoic acid structure is as follows:

[0010]

[0011] The lipoic acid selected in the present invention is a racemate, and its specific stereo conformation is not limited.

[0012] As a natural coenzyme, lipoic acid participates in the human body's metabolic process and has a 1,2-dithiolane ring, which has both biological safety and the potential for thiol exchange, solving the problem of high toxicity of current delivery carriers. The present invention uses it as a drug carrier to load chemotherapeutic drugs and / or photodynamic agents. The nano drug can deliver drugs through the thiol exchange pathway for single treatment or combined treatment of skin diseases, which is a new way that is different from the existing transdermal route.

[0013] Specifically, the present invention provides a nano drug transdermal delivery system, which uses lipoic acid as a drug carrier, encapsulates chemotherapeutic drugs and / or photodynamic agents, and forms nanoparticles containing 1,2-dithiolane through self-assembly.

[0014] The invention can encapsulate any one of chemotherapeutic drugs and photodynamic drugs to assemble into nanoparticles, or can encapsulate chemotherapeutic drugs and photodynamic agents to assemble into nanoparticles at the same time.

[0015] Optionally, the molar ratio of lipoic acid to chemotherapeutic drug or photodynamic agent is 5:1-50:1.

[0016] Optionally, the molar ratio of lipoic acid to the chemotherapeutic drug and the photodynamic agent is 5:1:1-50:1:1.

[0017] This ratio can achieve the best assembly effect.

[0018] Optionally, the chemotherapeutic drug is a hydrophobic chemotherapeutic drug, more preferably a first-line chemotherapeutic drug for the treatment of skin cancer, including but not limited to one or more of Trametinib, Dabrafenib, Paclitaxel, Doxorubicin, Docetaxel, Methotrexate, Vemurafenib, Cobimetinib, Gemcitabine, etc. The structural formula is shown below:

[0019]

[0020] Optionally, the photodynamic agent is a derivative of porphyrin, including but not limited to one or more of chlorin (Chlorine6), hematoporphyrin monomethyl ether, etc. The structural formula is as follows:

[0021]

[0022] The present invention continuously adjusts the molar ratio of lipoic acid to drug assembly according to the properties of the drug, and can obtain a variety of nano drugs with uniform particle sizes. Optionally, the particle size of the nanoparticles is 10-1000nm, preferably 50-500nm.

[0023] The present invention has no particular limitation on the ratio of the entrapped chemotherapeutic drugs and the photodynamic agent, which can be adjusted according to the focus and effect of the treatment. In some specific embodiments, the molar ratio of the chemotherapeutic drugs and the photodynamic agent is 1:1.

[0024] The present invention can realize the assembly with various drugs by adjusting the assembly molar number of lipoic acid, and can flexibly adjust the type of drug according to the target of treatment. The self-assembled nano drug shows tolerance to GSH concentration and time, which solves the problem that the current thiol exchange system is difficult to carry out multi-layer delivery due to being easily depolymerized by GSH. At the same time, the thiol exchange of the above-mentioned nano drug transdermal delivery system is carried out through dynamic disulfide exchange transdermal with the sulfhydryl group on the skin surface, that is, thiol exchange, and the transdermal efficiency is not affected by the physicochemical properties (size, shape, charge, etc.) of the nano drug itself, providing a new transdermal delivery route. The test results show that the above-mentioned nano drug transdermal delivery system provided by the present invention has excellent transdermal ability, which can reach subcutaneous>1200 microns. Thanks to the excellent transdermal ability, the above-mentioned nano drug transdermal delivery system achieves the complete eradication of melanoma, and the complete eradication of this disease can hardly be achieved by other methods except surgical excision.

[0025] The present invention also provides a method for preparing the nano drug transdermal delivery system, comprising the following steps:

[0026] The lipoic acid solution and the solution containing the chemotherapeutic drug and / or the photodynamic agent are mixed in water and self-assembled to obtain nanoparticles containing 1,2-dithiolane.

[0027] Optionally, the solvent of the lipoic acid solution is DMSO.

[0028] Optionally, in the solution containing the chemotherapeutic drug and / or photodynamic agent, the solvent is DMSO.

[0029] Optionally, the concentration of the lipoic acid solution is 74 μg / mL-740 μg / mL.

[0030] Optionally, in the solution containing chemotherapeutic drugs and / or photodynamic agents, the total concentration of chemotherapeutic drugs and / or photodynamic agents is 5.86 μg / mL-293 μg / mL.

[0031] The present invention also provides a transdermal nano drug composition, comprising the above nano drug transdermal delivery system and a pharmaceutically acceptable adjuvant.

[0032] The present invention has no particular limitation on the types of the above-mentioned adjuvants, which can be adjusted according to the dosage form.

[0033] The present invention has no particular limitation on the dosage form of the above-mentioned composition, and it can be a conventional dosage form well known in the art, including but not limited to gel, dressing, spray, patch, cream, etc.

[0034] The present invention provides the use of the nano drug transdermal delivery system or the transdermal nano drug composition in the preparation of drugs for preventing, alleviating or treating skin diseases.

[0035] The above-mentioned skin diseases include but are not limited to melanoma, psoriasis, basal carcinoma, squamous cell carcinoma, etc.

[0036] The nano drug transdermal delivery system or the transdermal nano drug composition provided by the present invention can deliver drugs that are difficult to penetrate into the subcutaneous layer through the skin surface in a simple, non-invasive local administration method (such as applying or spraying on the skin surface) to penetrate into the subcutaneous layer, reaching the lesions located in the dermis layer without the need for additional assistance. Not only is the administration convenient, but the toxic and side effects of chemotherapeutic drugs brought about by systemic administration can be reduced through transdermal administration. Nanoparticles can penetrate into the subcutaneous dermis layer to achieve complete elimination of subcutaneous tumors.

[0037] The nano drug transdermal delivery system or the transdermal nano drug composition can be used for single treatment or combined treatment of skin diseases.

[0038] In combination therapy, it can be used in combination with photodynamic agents.

[0039] In some specific embodiments, while applying the above-mentioned nano drug transdermal delivery system or the above-mentioned transdermal nano drug composition, appropriate laser irradiation is also performed on the affected area.

[0040] Compared with the prior art, the present invention provides a nano drug transdermal delivery system, which uses lipoic acid as a drug carrier, encapsulates chemotherapy drugs and / or photodynamic agents, and forms nanoparticles containing 1,2-dithiolane through self-assembly. The present invention is the first to use lipoic acid as a carrier-assisted small molecule in a transdermal delivery system.

[0041] In order to achieve efficient transdermal delivery through thiol exchange, nanomedicines need to break through the stratum corneum barrier and the cell barrier. There is a dynamic balance between thiol groups and disulfide bonds on the surface of the stratum corneum, which provides a theoretical basis for the application of the thiol exchange pathway to transdermal delivery. In the system that delivers drugs through the thiol exchange pathway, when taken up by cells, the disulfide bonds in its carrier will be reduced and depolymerized by glutathione (GSH) reductase in the cytoplasm, thereby releasing the loaded drugs to achieve the therapeutic purpose. However, after being depolymerized by GSH, the system also loses the ability to continue to deliver drugs to the next cell, which greatly limits the application of thiol exchange in drug delivery in multi-layer tissues. In particular, in some diseased tissues, such as tumor tissues, the concentration of GSH will increase significantly, and the depolymerization of the above-mentioned delivery system will also be accelerated. The present invention adopts lipoic acid as a carrier, and the prepared nano drug is incubated with GSH at a concentration of 10mM (the GSH concentration in general cancer tissue is 2-10mM, 2-10 times higher than normal tissue and above) for 4h before it is completely depolymerized, while the GSH concentration in normal tissue cells is generally 1-3mM. At this concentration, the nano drug provided by the present invention hardly depolymerizes. Therefore, the nano drug can maintain integrity during the process of penetrating the stratum corneum, epidermis and dermis, and realize the delivery of multi-layer skin tissue. When subcutaneous lesions occur, the GSH concentration of the disease site increases. At this time, the above-mentioned nano drug will continuously release the drug as time goes by after reaching the disease site through the stratum corneum. In this way, the penetration of multi-layer cells is achieved, overcoming the problem that the delivery drug can only reach a single layer of cells.

[0042] In summary, the present invention provides a transdermal delivery system containing 1,2-dithiolane, which efficiently delivers chemotherapeutic drugs and photodynamic agents through a thiol exchange pathway. Unlike other current transdermal pathways, the thiol exchange pathway performs dynamic disulfide exchange between cargo with disulfide bonds and thiol groups on the skin surface for transdermal delivery. No additional transdermal penetration enhancer is required, and it is not controlled by the position of the skin and the number of appendages. No equipment is required for auxiliary administration, and effective transdermal delivery can be achieved by simple application. The administration method is non-invasive, and efficient transdermal delivery can be achieved without destroying the stratum corneum, reducing the risk of skin infection. The transdermal depth can be more than 1 mm, which is much greater than the sum of the thickness of the human stratum corneum (thickness of about 18.4 microns) and the epidermis (about 49.5 microns), indicating that it is not only suitable for superficial skin diseases (disease sites in the epidermis), but also has potential therapeutic effects for skin diseases that occur in deep layers (disease sites in the dermis). It can be operated autonomously, significantly reducing costs. Lipoic acid is used as a carrier for auxiliary transdermal delivery in the present invention. It is a naturally occurring coenzyme that can be used for drug assembly without modification and has high safety. Based on this, the present invention can flexibly change the type of medicine to be loaded according to the type of skin disease to achieve more precise treatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0043] Figure 1 This is a characterization diagram of self-assembled nanoparticles of lipoic acid and chlorin e6;

[0044] Figure 2 Characterization diagram of self-assembled nanoparticles of lipoic acid and various chemotherapeutic drugs;

[0045] Figure 3 Characterization diagram of self-assembled nanoparticles of lipoic acid, various chemotherapeutic drugs, and photodynamic agents;

[0046] Figure 4 This is the tolerance result of Sup-TDDS (lipoic acid, trametinib and dihydrochlorin) to time at the same concentration of GSH;

[0047] Figure 5 This is a graph showing the tolerance of Sup-TDDS (lipoic acid, trametinib and chlorin assembly) to different GSH concentrations at the same time;

[0048] Figure 6 The result diagram of Sup-TDDS (lipoic acid, dabrafenib and chlorin assembly) effectively penetrating pig skin;

[0049] Figure 7 The result diagram of Sup-TDDS (lipoic acid, trametinib and chlorin assembly) effectively penetrating pig skin;

[0050] Figure 8 This is a result diagram showing that Sup-TDDS (lipoic acid, trametinib and hematoporphyrin monomethyl ether) effectively penetrates pig skin;

[0051] Fig. 9 This is a result diagram showing that Sup-TDDS (lipoic acid and chlorin assembly) effectively penetrates pig skin;

[0052] Fig.10 The result diagram of Sup-TDDS (lipoic acid, trametinib and chlorin assembly) effectively penetrating rat skin;

[0053] Fig.11 This is a study diagram of the transdermal mechanism of Sup-TDDS (lipoic acid, trametinib and dihydrochlorin assembly);

[0054] Fig.12 This is a diagram showing the results of tumor treatment after transdermal administration of Sup-TDDS (lipoic acid, trametinib and chlorin assembly);

[0055] Fig.13 The expression of phosphorylated extracellular regulated protein kinase (p-ERK) in tumor tissues after Sup-TDDS (lipoic acid, trametinib and chlorin assembly) transdermal treatment;

[0056] Fig.14 The expression of cadherin (CRT) and cytotoxic T cells (CD4+CD8+T cells) in tumor tissues and mature dendritic cells (CD80+CD86+) in tumor-draining lymph nodes after Sup-TDDS (lipoic acid, trametinib and dihydrochlorin assembly) transdermal treatment was completed;

[0057] Fig.15 This is a diagram showing the results of tumor treatment after transdermal administration of Sup-TDDS (lipoic acid, dabrafenib and dihydrochlorin assembly);

[0058] Fig.16 This is the hematoxylin and eosin (H&E) staining of mouse skin on the tenth day after the first transdermal administration of Sup-TDDS (lipoic acid, trametinib and dihydrochlorin assembly);

[0059] Fig.17 The results of tumor treatment after transdermal administration or tail vein administration of Sup-TDDS (lipoic acid, trametinib and dihydrochlorin assembly) are shown;

[0060] Fig.18 This is a graph showing the levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in serum after four days of transdermal or injection administration of Sup-TDDS (lipoic acid, trametinib and chlorin assembly) with a five-fold increase in drug dose;

[0061] Fig.19 The results of H&E sections of the liver after the drug dose of Sup-TDDS (lipoic acid, trametinib and dihydrochlorin assembly) was increased fivefold. DETAILED DESCRIPTION

[0062] In order to further illustrate the present invention, the nano drug transdermal delivery system provided by the present invention and its application in the treatment of skin diseases are described in detail below in conjunction with the embodiments. It should be understood that the above is only a specific embodiment of the present invention and is not intended to limit the present invention. Any modification, supplement and equivalent replacement made within the scope of the principles of the present invention should be included in the protection scope of the present invention.

[0063] In the following examples, raw materials and reagents not otherwise specified are all commercially available standard products.

[0064] Example 1 Assembly of lipoic acid and photodynamic reagent

[0065] Self-assembly of lipoic acid and chlorin

[0066] Ⅰ) Preparation of drug stock solution

[0067] Lipoic acid was prepared into a 50 mg / mL DMSO solution; dihydrochlorin was prepared into a 10 mg / mL DMSO solution;

[0068] II) Preparation of Nanoparticles

[0069] Take each component in (I) in a centrifuge tube according to the molar ratio (lipoic acid: dihydrochlorin) of 48:1, vortex and mix thoroughly. Add dropwise into deionized water while stirring, stir for 6 hours, dialyze for 48 hours, and centrifuge at 8000 rpm for 25 minutes to obtain the precipitate, which is the prepared nanoparticles with an average particle size of 142 nm. Figure 1 . Figure 1 Characterization diagram of self-assembled nanoparticles of lipoic acid and dihydrochlorin (Chlorin e6), where the left picture is the appearance photo of the assembled nanoparticles, the middle picture is the particle size distribution diagram of the nanoparticles obtained by dynamic light scattering particle size analyzer testing, and the right picture is the morphology of the nanoparticles taken under a transmission electron microscope.

[0070] Example 2 Assembly of lipoic acid and chemotherapeutic drugs

[0071] 2.1 Self-assembly of lipoic acid and cobimetinib

[0072] Ⅰ) Preparation of drug stock solution

[0073] Lipoic acid was prepared into a 50 mg / mL DMSO solution; cobimetinib was prepared into a 10 mM DMSO solution;

[0074] II) Preparation of Nanoparticles

[0075] Take the components in (I) at a molar ratio of 50:1 (lipoic acid: cobimetinib), add cobimetinib to water first, and mix thoroughly. Add lipoic acid solution dropwise while shaking on an oscillator, shake at 25°C and 750 rpm for 12 h to obtain the precipitate, which is the prepared nanoparticles; the average particle size is about 200 nm, such as Figure 2 As shown in Figure a, in Figure a, the left picture is a photo of the appearance of the assembled nanoparticles, the middle picture is a particle size distribution diagram of the nanoparticles obtained by testing with a dynamic light scattering particle size analyzer, and the right picture is a morphology of the nanoparticles taken under a transmission electron microscope.

[0076] 2.2 Self-assembly of lipoic acid and trametinib

[0077] Ⅰ) Preparation of drug stock solution

[0078] Lipoic acid was prepared into a 50 mg / mL DMSO solution; trametinib was prepared into a 10 mM DMSO solution;

[0079] II) Preparation of Nanoparticles

[0080] Take each component in (I) in a centrifuge tube at a molar ratio of 50:1 (lipoic acid: trametinib), vortex and mix thoroughly. Add dropwise into deionized water while stirring, stir for 6 hours, dialyze for 48 hours, and centrifuge at 8000 rpm for 25 minutes to obtain the prepared nanoparticles with an average particle size of 500 nm. Figure 2 As shown in Figure b, in Figure b, the left picture is a photo of the appearance of the assembled nanoparticles, the middle picture is a particle size distribution diagram of the nanoparticles obtained by testing with a dynamic light scattering particle size analyzer, and the right picture is a morphology of the nanoparticles taken under a transmission electron microscope.

[0081] 2.3 Self-assembly of lipoic acid and gemcitabine

[0082] Ⅰ) Preparation of drug stock solution

[0083] Lipoic acid: 50 mg / mL (DMSO solution)

[0084] Gemcitabine: 10 mg / mL (DMSO solution)

[0085] II) Preparation of Nanoparticles

[0086] The components in (I) were mixed in a centrifuge tube at a molar ratio of 35:1 (lipoic acid: gemcitabine) and then rapidly injected into vigorously stirred water to obtain nanoparticles with an average particle size of about 200 nm. Figure 2 As shown in Figure c, in Figure c, the left picture is a photo of the appearance of the assembled nanoparticles, the middle picture is a particle size distribution diagram of the nanoparticles obtained by testing with a dynamic light scattering particle size analyzer, and the right picture is a morphology of the nanoparticles taken under a transmission electron microscope.

[0087] 2.4 Self-assembly of lipoic acid and methotrexate

[0088] Ⅰ) Preparation of drug stock solution

[0089] Lipoic acid was prepared into a 50 mg / mL DMSO solution; methotrexate was prepared into a 10 mM DMSO solution;

[0090] II) Preparation of Nanoparticles

[0091] Take the components in (I) according to the molar ratio (lipoic acid: methotrexate) of 50:1, add methotrexate to water first, and mix thoroughly. Add lipoic acid solution dropwise while shaking on an oscillator, shake at 25°C and 750 rpm for 12 hours, and the resulting precipitate is the prepared nanoparticles with an average particle size of about 200 nm. Figure 2 As shown in Figure d, in Figure d, the left picture is a photo of the appearance of the assembled nanoparticles, the middle picture is a particle size distribution diagram of the nanoparticles obtained by testing with a dynamic light scattering particle size analyzer, and the right picture is a morphology of the nanoparticles taken under a transmission electron microscope.

[0092] Examples 1 and 2 show that lipoic acid can be assembled into nanoparticles with chemotherapeutic drugs or photodynamic agents.

[0093] Example 3 Self-assembly of lipoic acid, chemotherapeutic drugs and photodynamic agents

[0094] 3.1 Self-assembly of lipoic acid, dabrafenib and chlorin

[0095] Ⅰ) Preparation of drug stock solution

[0096] Lipoic acid was prepared into a 50 mg / mL DMSO solution; dabrafenib was prepared into a 10 mg / mL DMSO solution; and chlorin was prepared into a 10 mg / mL DMSO solution.

[0097] II) Preparation of Nanoparticles

[0098] Take the stock solution of each component in (I) in a centrifuge tube according to the molar ratio (lipoic acid: dabrafenib: dihydrochlorin) of 5:1:1, vortex and mix thoroughly. Add dropwise into deionized water while stirring, stir for 6 hours, dialyze for 48 hours, centrifuge at 8000 rpm for 25 minutes, and the obtained precipitate is the prepared nanoparticles. The average particle size is 144nm. Figure 3 As shown in Figure a, in Figure a, the left picture is a photo of the appearance of the assembled nanoparticles, the middle picture is a particle size distribution diagram of the nanoparticles obtained by testing with a dynamic light scattering particle size analyzer, and the right picture is a morphology of the nanoparticles taken under a transmission electron microscope.

[0099] 3.2 Self-assembly of lipoic acid, trametinib and chlorin

[0100] Ⅰ) Preparation of drug stock solution

[0101] Lipoic acid was prepared into a 50 mg / mL DMSO solution; trametinib was prepared into a 10 mM DMSO solution; and chlorin was prepared into a 10 mg / mL DMSO solution.

[0102] II) Preparation of Nanoparticles

[0103] According to the molar ratio of (lipoic acid: trametinib: dihydrochlorin) of 14:1:2, the stock solution of each component in (I) was taken into a centrifuge tube, vortexed and mixed thoroughly. It was added dropwise into deionized water while stirring, stirred for 6 hours, dialyzed for 48 hours, and centrifuged at 8000 rpm for 25 minutes. The precipitate obtained was the prepared nanoparticles with an average particle size of 163nm. Figure 3 As shown in Figure b, in Figure b, the left picture is a photo of the appearance of the assembled nanoparticles, the middle picture is a particle size distribution diagram of the nanoparticles obtained by testing with a dynamic light scattering particle size analyzer, and the right picture is a morphology of the nanoparticles taken under a transmission electron microscope.

[0104] 3.3 Self-assembly of lipoic acid, vemurafenib and chlorin

[0105] Ⅰ) Preparation of drug stock solution

[0106] Lipoic acid was prepared into a 50 mg / mL DMSO solution; Vemurafenib was prepared into a 10 mM DMSO solution; and chlorin was prepared into a 10 mg / mL DMSO solution.

[0107] II) Preparation of Nanoparticles

[0108] Take the stock solution of each component in (I) in a centrifuge tube according to the molar ratio (lipoic acid: vemurafenib: dihydrochlorin) of 14:1:2, vortex and mix thoroughly. Add dropwise into deionized water while stirring, stir for 6 hours, dialyze for 48 hours, centrifuge at 8000 rpm for 25 minutes, and the obtained precipitate is the prepared nanoparticles with an average particle size of 132nm. Figure 3 As shown in Figure c, in Figure c, the left picture is a photo of the appearance of the assembled nanoparticles, the middle picture is a particle size distribution diagram of the nanoparticles obtained by testing with a dynamic light scattering particle size analyzer, and the right picture is a morphology of the nanoparticles taken under a transmission electron microscope.

[0109] 3.4 Self-assembly of lipoic acid, docetaxel and chlorin

[0110] Ⅰ) Preparation of drug stock solution

[0111] Lipoic acid was prepared into a 50 mg / mL DMSO solution; docetaxel was prepared into a 10 mM DMSO solution; and chlorin was prepared into a 10 mg / mL DMSO solution.

[0112] II) Preparation of Nanoparticles

[0113] According to the molar ratio (lipoic acid: docetaxel: dihydrochlorin) of 14:1:2, the stock solution of each component in (I) was taken into a centrifuge tube, vortexed, and mixed thoroughly. It was added dropwise into deionized water while stirring, stirred for 6 hours, dialyzed for 48 hours, and centrifuged at 8000 rpm for 25 minutes. The precipitate obtained was the prepared nanoparticles with an average particle size of 148nm. Figure 3 As shown in Figure d, in Figure d, the left picture is a photo of the appearance of the assembled nanoparticles, the middle picture is a particle size distribution diagram of the nanoparticles obtained by testing with a dynamic light scattering particle size analyzer, and the right picture is a morphology of the nanoparticles taken under a transmission electron microscope.

[0114] 3.5 Self-assembly of lipoic acid, paclitaxel and chlorin

[0115] Ⅰ) Preparation of drug stock solution

[0116] Lipoic acid was prepared into a 50 mg / mL DMSO solution; paclitaxel was prepared into a 10 mM DMSO solution; and chlorin was prepared into a 10 mg / mL DMSO solution.

[0117] II) Preparation of Nanoparticles

[0118] Take each component in (I) in a centrifuge tube according to the molar ratio (lipoic acid: paclitaxel: dihydrochlorin) of 14:1:2, vortex and mix thoroughly. Add dropwise into deionized water while stirring, stir for 6 hours, dialyze for 48 hours, centrifuge at 8000 rpm for 25 minutes, and the obtained precipitate is the prepared nanoparticles with an average particle size of 142nm. Figure 3 As shown in Figure e, in Figure e, the left picture is a photo of the appearance of the assembled nanoparticles, the middle picture is a particle size distribution diagram of the nanoparticles obtained by testing with a dynamic light scattering particle size analyzer, and the right picture is a morphology of the nanoparticles taken under a transmission electron microscope.

[0119] 3.6 Self-assembly of lipoic acid, doxorubicin and chlorin e6

[0120] Ⅰ) Preparation of drug stock solution

[0121] Lipoic acid was prepared into a 50 mg / mL DMSO solution; doxorubicin was prepared into a 10 mg / mL DMSO solution; and chlorin was prepared into a 10 mg / mL DMSO solution.

[0122] II) Preparation of Nanoparticles

[0123] Take the components in (I) in a centrifuge tube according to the molar ratio of lipoic acid: doxorubicin: hematoporphyrin monomethylether (Hematoporphyrin monomethylether) of 29:1:1, vortex and mix thoroughly. Add dropwise into deionized water while stirring, stir for 6 hours, dialyze for 48 hours, centrifuge at 8000 rpm for 25 minutes, and the obtained precipitate is the prepared nanoparticles with an average particle size of 807 nm. Figure 3 As shown in Figure f, in Figure f, the left picture is a photo of the appearance of the assembled nanoparticles, the middle picture is a particle size distribution diagram of the nanoparticles obtained by testing with a dynamic light scattering particle size analyzer, and the right picture is a morphology of the nanoparticles taken under a transmission electron microscope.

[0124] 3.7 Self-assembly of lipoic acid, trametinib and hematoporphyrin monomethyl ether

[0125] Ⅰ) Preparation of drug stock solution

[0126] Lipoic acid was prepared into a 50 mg / mL DMSO solution; trametinib was prepared into a 10 mM DMSO solution; and hematoporphyrin monomethyl ether was prepared into a 10 mM DMSO solution.

[0127] II) Preparation of Nanoparticles

[0128] Take the components in (I) in a centrifuge tube according to the molar ratio (lipoic acid: trametinib: hematoporphyrin monomethyl ether) of 48:1:1, vortex and mix thoroughly. Add dropwise into deionized water while stirring, stir for 6 hours, dialyze for 48 hours, centrifuge at 8000 rpm for 25 minutes, and the obtained precipitate is the prepared nanoparticles with an average particle size of 142 nm. Figure 3 As shown in Figure g, in Figure g, the left picture is a photo of the appearance of the assembled nanoparticles, the middle picture is a particle size distribution diagram of the nanoparticles obtained by testing with a dynamic light scattering particle size analyzer, and the right picture is a morphology of the nanoparticles taken under a transmission electron microscope.

[0129] Example 3 shows that lipoic acid can be assembled into nanoparticles together with various chemotherapeutic drugs and photodynamic agents.

[0130] Example 4: Nanoparticles assembled from lipoic acid and multiple drugs have GSH tolerance

[0131] Ⅰ) Preparation of 100 mM GSH aqueous solution

[0132] Ⅱ) Nanoparticles and 10 mM GSH were incubated for different time periods

[0133] Four tubes of the nanoparticles in Example 3.2 containing 10 mM GSH were placed in a shaker at 37° C. and incubated for 0.5 h, 1 h, 2 h, and 4 h, respectively, and then photographed for appearance.

[0134] Since chemotherapy drugs and dihydrochlorins are extremely hydrophobic, the five-membered disulfide ring of lipoic acid in the nanomedicine is gradually reduced over time, the nanomedicine gradually depolymerizes and releases the loaded drugs, and the precipitated free drugs aggregate and precipitate in water due to their strong hydrophobicity.

[0135] The test results are shown in Figure 4 It can be seen that 10mM GSH completely depolymerized the nanodrug and released the free drug only after incubation with the nanodrug for 4h.

[0136] III) Preparation of nanoparticles containing different concentrations of GSH

[0137] The total volume was 200 μL, and the nanoparticles were prepared with concentrations of 0.1, 0.2, 0.5, 1, 2, 4, 5, 8, and 10 mM GSH. Taking 10 mM as an example, 20 μL of solution I) was taken into a 1.5 mL centrifuge tube, and 180 μL of the nanoparticles in Example 3.2 was added and mixed. The preparations of other concentrations were similar, and the appearance was photographed after incubation in a shaker at 37°C for 4 hours.

[0138] The test results are shown in Figure 5 , it can be seen that GSH with a concentration lower than 4 mM hardly affects the integrity of the nanodrug even after being incubated with the nanodrug for 4 h.

[0139] Example 5 Lipoic acid and multiple drugs assembled into nanoparticles have transdermal ability

[0140] 5.1 Nanoparticles assembled with lipoic acid, dabrafenib and chlorin as a specific embodiment for transdermal study

[0141] Ⅰ) Nanoparticle preparation is the same as above

[0142] II) Quantification of Chlorins in Nanoparticles

[0143] Weigh the standard chlorin to prepare concentrations of 6.25 μg mL -1 ,12.5μg mL -1 ,50μgmL -1 ,100μgmL -1 ,200μg mL -1 The DMSO solution was used as the standard solution, and the absorption value at 660 nm was measured by UV-visible absorption spectrometer. The concentration of dihydrochlorin was taken as X value, and the measured absorption value was taken as Y value. The standard curve of dihydrochlorin was obtained as Y=0.005X-0.0058, R 2 =0.9995.

[0144] 5 μL of the purified nanoparticles were taken and depolymerized with 195 μL of DMSO, and the absorption value at 660 nm was measured using a UV-visible absorption spectrometer. After being inserted into the standard curve, the concentration of dihydrochlorin in the nanoparticles was calculated.

[0145] III) Penetration effect in pig skin

[0146] Fresh pig skin was selected from the local market. Subcutaneous fat was carefully removed, and the pig skin was cut into 1 cm squares and a circle was drawn with a radius of 0.5 mm. A small amount of nanoparticle aqueous solution containing 50 μg of dihydrochlorin was added to the circle marked on the pig skin several times. After incubation for 10 min, 60 min and 240 min, the pig skin was scrubbed with PBS three times, frozen and cut into 10 μm slices and placed on a slide. DAPI staining solution was used to mark the cell nucleus, and the penetration depth of the drug in the skin was observed by laser confocal microscopy. Among them, dihydrochlorin aqueous solution and nanoparticles assembled with dihydrochlorin and chemotherapeutic drugs were used as controls. Dihydrochlorin aqueous solution group (free Ce6), nanoparticle group assembled with dihydrochlorin and chemotherapeutic drugs (Inf-TDDS), lipoic acid, chemotherapeutic drug dabrafenib and nanoparticle group assembled with dihydrochlorin (Sup-TDDS).

[0147] The results are as follows Figure 6 .Depend on Figure 6 It can be seen that Sup-TDDS can effectively penetrate the stratum corneum. As the incubation time increases, the amount of drug penetration increases and the penetration becomes deeper. After 4 hours of administration, it can penetrate more than 600 microns into the skin, while free Ce6 and Inf-TDDS can hardly penetrate the stratum corneum. This shows that the transdermal drug delivery system of the present invention has excellent transdermal ability.

[0148] 5.2 Nanoparticles assembled with lipoic acid, trametinib and chlorin as a specific embodiment for transdermal study

[0149] The operation steps are the same as 5.1. The experimental results are as follows Figure 7 The test results and Figure 6 Consistent.

[0150] 5.3 Nanoparticles assembled with lipoic acid, trametinib and hematoporphyrin monomethyl ether as a specific embodiment for transdermal study

[0151] The operation steps are the same as in Example 5.1, and the experimental results are as follows Figure 8 , Figure 8 In the figure, the left picture shows the fluorescence imaging results of hematoporphyrin monomethyl ether (red) in the nanoparticles assembled by trametinib and hematoporphyrin monomethyl ether in pig skin and the merged image of the cell nucleus (blue) in pig skin. The right picture shows the single channel result of hematoporphyrin monomethyl ether (red) in the merged channel of the left picture. Figure 6 Consistent.

[0152] 5.4 Nanoparticles assembled with lipoic acid and chlorin as a specific embodiment for transdermal studies

[0153] The operation steps are the same as in Example 5.1, and the experimental results are as follows Fig. 9 , Fig. 9 In the figure, the left picture shows the fluorescence imaging results of dihydrochlorin (red) in the nanoparticles assembled by lipoic acid and dihydrochlorin in pig skin and the merged image of the cell nucleus (blue) in pig skin. The right picture shows the single channel result of dihydrochlorin (red) in the merged channel of the left picture. Figure 6 Consistent.

[0154] The above results show that the transdermal ability of the assembly containing lipoic acid is significantly higher than that of free drugs and nanoparticles assembled from free drugs and photodynamic agents, which can indicate that lipoic acid can significantly improve the transdermal ability of nanodrugs. By adjusting the molar ratio of lipoic acid to small molecule drugs, the penetration depth of nanoparticles increased from more than 600 microns to more than 1200 microns. This shows that by adjusting the assembly ratio of lipoic acid, the penetration depth of nanodrugs can be regulated. Therefore, the proportion of lipoic acid can be adjusted according to the disease site in the skin for precise disease treatment.

[0155] Example 6 Lipoic acid and multiple drugs assembled into nanoparticles have the ability to penetrate the skin in vivo

[0156] This example uses the lipoic acid, trametinib and dihydrochlorin to assemble nanoparticles as a specific example to verify its transdermal effect on living rats.

[0157] Ⅰ) Rats stable period

[0158] Female SD rats weighing 200 g were purchased and housed in an animal room for one week.

[0159] II) Transdermal operation in living rats

[0160] The rats were anesthetized, the hair on the back of the rats was carefully removed, and a circle with a radius of 0.5 mm was drawn on the back. A small amount of an aqueous solution of nanoparticles containing 50 μg of chlorin e6 was added to the circle on the back several times. After incubation for 1 h, 4 h and 12 h, the administration site was scrubbed three times with PBS (phosphate buffered saline). After the rats were euthanized, the skin at the administration site was peeled off, frozen and cut into 10-micron slices and placed on a glass slide. The cell nuclei were marked with DAPI staining solution, and the penetration depth of the drug in the skin was observed by laser confocal microscopy. Among them, the chlorin aqueous solution and the nanoparticles assembled with chlorin and the chemotherapy drug trametinib were used as controls.

[0161] The results are as follows Fig.10 . Fig.10 Aqueous solution of chlorin (free Ce6), nanoparticles assembled from chlorin and trametinib (Inf-TDDS), nanoparticles assembled from lipoic acid, trametinib and chlorin (Sup-TDDS). As shown in the figure, Sup-TDDS can effectively penetrate the stratum corneum. As the incubation time increases, the amount of drug penetration increases and the penetration becomes deeper. After 12 hours of administration, it can penetrate more than 600 microns into the skin, while free Ce6 and Inf-TDDS can hardly penetrate the stratum corneum. This shows that the transdermal drug delivery system of the present invention can be applied to living bodies.

[0162] Example 7 Mechanism of transdermal penetration of nanoparticles assembled with lipoic acid and multiple drugs

[0163] Nanoparticles assembled with lipoic acid, trametinib and dihydrochlorin are used as a specific embodiment for the study of transdermal treatment of skin melanoma.

[0164] Implementation steps:

[0165] Ⅰ) Nanoparticle preparation and quantification are the same as above

[0166] II) Quantification of chlorin in nanoparticles Same as above

[0167] III) Penetration mechanism in pig skin

[0168] The pig skin treatment and administration method were the same as in Example 6, except that, before the administration and incubation, DTNB (5,5' dithiobis(2-nitrobenzoic acid) as a thiol exchange inhibitor) was first fully applied to the pig skin surface and incubated with the pig skin for 30 minutes.

[0169] The experimental results are shown in Fig.11 , Fig.11 Aqueous solution of chlorin (free Ce6), nanoparticles assembled from chlorin and trametinib (Inf-TDDS), nanoparticles assembled from lipoic acid, trametinib and chlorin (Sup-TDDS). As shown in the figure, after DTNB pretreatment of the skin, Sup-TDDS cannot effectively penetrate the stratum corneum, while the penetration effects of free Ce6 and Inf-TDDS are not affected by DTNB. It can be seen that DTNB can completely inhibit the transdermal penetration of nanomedicines, indicating that the transdermal system of the present invention is mainly carried out through the thiol exchange pathway.

[0170] The ability of lipoic acid, chemotherapeutic drugs and photodynamic agents assembled into nanoparticles to treat skin melanoma via transdermal administration as described in Example 8

[0171] Nanoparticles assembled with lipoic acid, trametinib and dihydrochlorin are used as a specific embodiment for the study of transdermal treatment of skin melanoma.

[0172] Implementation steps:

[0173] Ⅰ) Nanoparticle preparation and quantification are the same as above

[0174] II) Construction and grouping of skin melanoma mouse model

[0175] Purchase 30 C57 BL / 6 female mice weighing 20 g, 4-6 weeks old. 1x 10 6 B16F10 cells were inoculated into the right thigh of the mouse, and tumors with a size of 50-70 mm were obtained ten days later. 3 Skin melanoma mouse model. The mice were randomly divided into six groups, 5 mice in each group. PBS group, laser irradiation group only (Laser), chlorin aqueous solution and laser irradiation group (FreeCe6(+)), chlorin and trametinib assembled nanoparticles and laser irradiation group (Ihf-TDDS(+)), lipoic acid, chlorin and trametinib assembled nanoparticles without laser irradiation group (Sup-TDDS(-)), lipoic acid, chlorin and trametinib assembled nanoparticles and laser irradiation group (Sup-TDDS(+)).

[0176] III) Transdermal drug delivery for melanoma

[0177] Nanoparticles containing 50 μg of Chlorin e6 were added to the tumor site of mice in small amounts and multiple times. After 12 hours of administration, the tumor site was irradiated with a 660nm laser at a laser intensity of 0.5W for 10 minutes. The drug was administered three times in total, with one administration and laser irradiation every other day. The PBS group, the nanoparticles assembled with lipoic acid, Chlorin e6 and trametinib were not irradiated with laser. From the first administration, the tumor volume of the mice was observed and recorded (V = a × b 2 / 2; a: tumor length; b: tumor width), body weight, and survival time. Among them, the tumor volume of mice was greater than 2000mm 3 The mice were considered dead. The other groups were operated in parallel. Fig.12 . Fig.12 Figure a is a schematic diagram of the treatment method, Figure b is a line graph of tumor growth in mice in different dosing groups, Figure c is a line graph of tumor growth for each mouse in different dosing groups, Figure d is a statistical graph of survival time of mice in different dosing groups, and Figure e is a photo of completely cured mouse tumors in the Sup-TDDS laser irradiation group.

[0178] Fig.12 It can be seen that compared with PBS, laser, free Ce6(+), Inf-TDDS(+), and Sup-TDDS(-), Sup-TDDS(+) can achieve complete elimination of tumors and greatly prolong the survival time of mice. The above results show that the excellent transdermal ability of the transdermal system of the present invention can significantly improve the therapeutic effect, and due to the deep penetration ability, complete eradication of tumors is achieved.

[0179] IV) Verification of the effect of trametinib in Sup-TDDS

[0180] After the treatment cycle, the mouse tumor tissues were taken for immunohistochemical staining analysis. The mouse tissues were fixed with 4% paraformaldehyde, dehydrated with 30% sucrose, embedded with embedding medium, and cut into 10 μm tissue sections with a freezing microtome for immunohistochemical operation.

[0181] Fig.13 It can be seen that compared with the groups without lipoic acid components such as PBS, laser, free Ce6(+), and Inf-TDDS(+), the phosphorylated extracellular regulated protein kinase (p-ERK) in the tumor site was significantly downregulated in the Sup-TDDS(-) and Sup-TDDS(+) groups due to the effect of lipoic acid-assisted drug transdermal delivery. This is the pathway protein inhibited by the trametinib inhibitor, indicating that trametinib can be successfully delivered to tissues and exert an inhibitory effect.

[0182] V) Verify the immune effect induced by photodynamic therapy of chlorin e6 in Sup-TDDS

[0183] Tumor tissue and tumor-draining lymph nodes of mice at the end of the treatment cycle were taken for immunohistochemical staining and immune cell analysis. A portion of the mouse tumor tissue was fixed with 4% paraformaldehyde, dehydrated with 30% sucrose, embedded in an embedding agent, cut into 10μm tissue sections with a freezing microtome, and subjected to immunohistochemical fluorescent staining. A portion of the tumor tissue was dispersed into a single cell suspension by mechanical grinding and sieving, and then the cells were counted. One million cells were taken from each group for immunostaining and analyzed on the machine. Tumor-draining lymph nodes were immunostained in the same way.

[0184] Depend on Fig.14 As shown in Figure a, compared with PBS, laser, free Ce6(+), Inf-TDDS(+), and Sup-TDDS(-), the expression of cadherin (CRT) in the tumor tissue of the Sup-TDDS(+) group was significantly upregulated, which is a marker of immunogenic cell death. Its upregulation indicates improved tumor immunity. Its significant upregulation in the Sup-TDDS(+) group indicates that the dihydrochlorin (Chlorin e6) delivered to the tumor tissue caused apoptosis of tumor cells under laser irradiation and may induce an immune response. Fig.14 Figure b shows the immune analysis of the tumor-draining lymph nodes. It can be seen that the expression of dendritic cell maturation markers CD80 and CD86 was significantly increased in the Sup-TDDS(+) group, indicating that an immune response was induced. Fig.14 Figure c shows further immune cell analysis of tumor tissue, and the results show that the expression of cytotoxic T cell markers CD4 and CD8 was significantly increased in the Sup-TDDS(+) group, indicating that photodynamic therapy successfully induced an immune response.

[0185] VI) Research on the transdermal treatment of skin melanoma using nanoparticles assembled from lipoic acid, dabrafenib and dihydrochlorin as a specific embodiment. The implementation steps are the same as those in the steps of using nanoparticles assembled from lipoic acid, trametinib and dihydrochlorin for the treatment of melanoma, Ⅰ), Ⅱ) and Ⅲ)

[0186] The results are as follows Fig.15 As shown. Fig.15 It can be seen that compared with PBS, laser, free Ce6(+), Inf-TDDS(+), and Sup-TDDS(-), Sup-TDDS(+) can achieve complete elimination of tumors, indicating that the transdermal delivery system of the present invention can be assembled with different drugs and produce therapeutic effects.

[0187] Example 9 Verification of whether skin is damaged after transdermal drug administration

[0188] The specific implementation method is the same as that of Example 8. Ten days after the first administration, the cured mice in the Sup-TDDS(+) group were killed, the skin of the laser irradiated area was peeled off, cut into 10 μm slices and stained with H&E, and the tissue was observed and photographed under an inverted optical microscope. The skin of healthy mice without tumor grafting was used as a control. The results are shown in Figure 2. Fig.16 .

[0189] like Fig.16 As shown, compared with healthy mice without tumor implantation, the mouse skin after laser irradiation has no obvious morphological changes, indicating that the transdermal drug delivery system of the present invention will not cause skin damage and is non-invasive, indicating that the laser intensity used in the present invention will not cause skin trauma.

[0190] Example 10 Comparison of the effects of transdermal administration and traditional intravenous administration in the treatment of skin melanoma

[0191] The nanoparticles assembled from lipoic acid, trametinib and chlorin are used as a specific example in this embodiment.

[0192] Specific implementation steps:

[0193] Ⅰ) Preparation and quantification of nanoparticles and establishment of skin melanoma mouse model were the same as above

[0194] II) Comparison of the efficacy of transdermal administration and tail vein injection in the treatment of cutaneous melanoma

[0195] Take nanoparticles containing 50μg of Chlorin e6 and drip them into the tumor site of mice in small amounts and multiple times. Take the same amount of medicine and inject it into the tail vein. 12 hours after the administration, use a 660nm laser with a laser intensity of 0.5W to irradiate the tumor site for 10 minutes. The drug is administered three times, with an interval of one day between administration and laser irradiation. Starting from the first administration, observe and record the tumor volume of the mice (V = a × b 2 / 2; a: tumor length; b: tumor width), body weight. Among them, the tumor volume of mice was greater than 2000mm 3 The mice were considered dead. Fig.17 . Fig.17 In the figure, Figure a is a schematic diagram of the tail vein injection and transdermal administration methods, Figure b is a line graph of mouse tumor growth under two different administration methods, Figure c is a line graph of the weight change of mice during the treatment period, and Figure d is a line graph of the growth of each mouse tumor under two different administration methods.

[0196] like Fig.17As shown, tail vein injection only inhibited the growth of tumors in the early stage of administration, but in the later stage of treatment, the tumor grew rapidly again, indicating that tail vein injection cannot effectively eradicate tumors. In contrast, in the transdermal administration group, the mouse tumors were completely eradicated and there was no recurrence after drug withdrawal, indicating that the transdermal drug delivery system of the present invention greatly improved the cure rate of skin melanoma. The above results show that since transdermal administration can achieve a large amount of drug enrichment in the tumor, complete eradication of the tumor is achieved and the tumor does not recur. However, the drug administration method of this system, tail vein injection, leads to incomplete tumor treatment and easy tumor recurrence due to insufficient drug accumulation in the tumor.

[0197] Example 11 Comparison of the safety of transdermal administration and traditional intravenous administration after the dosage was increased five times compared with the above dosage

[0198] Specific implementation steps:

[0199] Ⅰ) Preparation and quantification of nanoparticles and establishment of skin melanoma mouse model were the same as above.

[0200] Ⅱ) Compare the safety of transdermal administration and tail vein injection.

[0201] Nanoparticles containing 250 μg of dihydrochlorin were dripped into the tumor site of mice in small amounts and multiple times. The same amount of drug was injected into the tail vein. The behavior of the mice was observed, and the mice were killed on the fourth day of administration. Blood was taken for liver function evaluation. The liver tissue was dissected, sliced, and stained with hematoxylin and eosin to observe the morphology of the liver tissue. The results are as follows: Fig.16 and Fig.17 .

[0202] Fig.18 Figure a shows the values ​​of alanine aminotransferase (ALT) in mouse serum under two different administration methods after the dosage was increased by 5 times, and Figure b shows the values ​​of aspartate aminotransferase (AST) in mouse serum under two different administration methods. Fig.18 As shown, in the tail vein injection group, the levels of ALT and AST increased significantly, even exceeding the upper limit of normal values, indicating that the tail vein injection of large doses of drugs will produce systemic toxicity, which is not conducive to the treatment of large tumors that require higher doses. In the transdermal administration group, the levels of ALT and AST were at normal levels, indicating that the transdermal drug delivery system of the present invention has great biosafety.

[0203] like Fig.19 As shown, in the tail vein injection group, multiple inflammatory cell infiltrations appeared in the liver tissue (indicated by the black arrows), indicating that the tail vein injection damaged the liver of the mice. In contrast, in the transdermal administration group, there was no morphological change in the mouse liver and no inflammatory cell infiltration, indicating that the transdermal administration of the present invention has a high safety.

[0204] The above results show that transdermal administration does not cause systemic toxicity because less of the drug enters the systemic circulation. Large-dose transdermal administration can be used to treat large tumors that require higher doses of drugs.

[0205] The above embodiments are only used to help understand the method and core idea of ​​the present invention. It should be noted that, for those skilled in the art, several improvements and modifications can be made to the present invention without departing from the principles of the present invention, and these improvements and modifications also fall within the scope of protection of the claims of the present invention.

Claims

1. Application of lipoic acid as a transdermal nanodrug carrier.

2. A nano drug transdermal delivery system, characterized in that: Lipoic acid is used as a drug carrier to encapsulate chemotherapy drugs and / or photodynamic agents, and nanoparticles containing 1,2-dithiolane are formed through self-assembly.

3. The nano drug transdermal delivery system according to claim 1, characterized in that: The molar ratio of lipoic acid to chemotherapeutic drugs or photodynamic agents is 5:1-50:1; The molar ratio of lipoic acid to the chemotherapy drug and the photodynamic agent is 5:1:1-50:1:

1.

4. The nano drug transdermal delivery system according to claim 1, characterized in that: The chemotherapy drug is selected from one or more of trametinib, dabrafenib, paclitaxel, doxorubicin, docetaxel, methotrexate, vemurafenib, and gemcitabine; The photodynamic agent is selected from one or more of dihydrochlorin and hematoporphyrin monomethyl ether.

5. The nano drug transdermal delivery system according to claim 1, characterized in that: The particle size of the nanoparticles is 10-1000 nm.

6. The method for preparing the nano drug transdermal delivery system according to any one of claims 2 to 5, comprising the following steps: The lipoic acid solution and the solution containing the chemotherapeutic drug and / or the photodynamic agent are mixed in water and self-assembled to obtain nanoparticles containing disulfide bonds.

7. The preparation method according to claim 6, characterized in that: The solvent of the lipoic acid solution is DMSO; In the solution containing the chemotherapeutic drug and / or photodynamic agent, the solvent is DMSO.

8. The preparation method according to claim 6, characterized in that: The concentration of the lipoic acid solution is 74 μg / mL-740 μg / mL; In the solution containing the chemotherapeutic drug and / or the photodynamic agent, the total concentration of the chemotherapeutic drug and / or the photodynamic agent is 5.86 μg / mL-293 μg / mL.

9. A transdermal nano drug composition, comprising the nano drug transdermal delivery system according to any one of claims 2 to 5 and a pharmaceutically acceptable adjuvant.

10. Use of the nano drug transdermal delivery system according to any one of claims 2 to 5 or the transdermal nano drug composition according to claim 9 in the preparation of drugs for preventing, alleviating or treating skin diseases.

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